Target genome engineering is desirable for many scientists. By deleting or inserting a designed and specific nucleotide sequence in an endogenous genome, scientists can generate various animal models for performing fundamental biological research and studying mechanisms of disease. In addition, scientists can create transgenic animals to produce biological compositions and/or components, which may be difficult to obtain from other resources. However, it is challenging to perform targeted and specific genome modifications using traditional techniques. The traditional techniques rely on random fragment exchanges of homologous chromosomes in natural cellular processes. Therefore, the efficiency for the traditional techniques is low (e.g., 10−6-10−8 as a successfully rate). Because of this low efficiency, these techniques are generally applied in mice rather than other animal models (e.g., large mammalians).
In 2009, two research groups identified a transcription activator-like effector (TALE) in plant pathogen Xanthomonas, which modulates host gene functions by binding specific sequences within gene promoters. The TALE related techniques helped scientists develop an easier method for targeted genome engineering. This technique fuses TALE to Fokl to generate a transcription activator-like effector nuclease (TALEN). In general, TALEs include tandem-like and nearly identical monomers (i.e., repeat domains), flanked by N-terminal and C-terminal sequences. Each monomer contains 34 amino acids, and the sequence of each monomer is highly conserved. Only two amino acids per repeat (i.e., residues 12th and 13th) are hypervariable, and are also known as repeat variable di-residues (RVDs). The RVDs determines the nucleotide-binding specificity of each TALE repeat domain.
TALE related techniques have increased the efficiency and usages of genome engineering, and make the genome engineering more convenient. However, assembling ten to twenty highly conserved DNA modules into a vector is a big challenge.